Toll-like receptors (TLRs) are a family of transmembrane proteins that recognize conserved microbial molecules, referred to as pathogen-associated molecular patterns, which are distinguishable from host molecules. As such TLRs play important roles in innate immune responses. TLR3, TLR7, TLR8, TLR9 and TLR13 are nucleic acid sensing TLRs.
Agonists and antagonists of TLRs find use in modulating immune responses. TLR agonists are typically employed to stimulate immune responses, whereas TLR antagonists are typically employed to inhibit immune responses (Gosu et al., Molecules, 17:13503-13529, 2012). TLR9, which is expressed by various antigen presenting cells, recognizes unmethylated CpG dinucleotides within nucleic acids. Thus polynucleotides containing an unmethylated CpG dinucleotide can make effective adjuvants through their ability to activate TLR9. Additionally, chimeric compounds containing both a non-nucleic acid moiety and an unmethylated-CpG containing nucleic acid moiety are capable of stimulating immune responses.
The potency of a TLR9 agonist is dependent upon the length of the nucleic acid moiety, the residues flanking the unmethylated-CpG dinucleotide, and the efficacy of antigen presenting cell uptake (Marshall et al., Nucleic Acids Research, 31:5122-5133, 2003). Certain branched chimeric compounds, in which multiple polynucleotides or linear chimeric compounds are attached to a multivalent carrier moiety (e.g., a polysaccharide), have elicited enhanced immune responses relative to the unconjugated polynucleotides or linear chimeric compounds (Marshall et al., supra). A highly branched hydrophilic polysaccharide, marketed as FICOLL® by GE Healthcare, can be used as a multivalent carrier moiety for branched chimeric compounds. However, traditional linker moieties used in conjugation of FICOLL® are hydrophobic, which can cause precipitation of the synthetic intermediates containing FICOLL®. This negatively impacts the processes used to manufacture the branched chimeric compounds and subsequent ability to store the final products. Moreover, the therapeutic utility of a synthetic TLR agonist is influenced by its toxicity.
There remains a need for polynucleotides and chimeric compounds with potent immunostimulatory activity. Additionally, there remains a need for potent chimeric compounds that can be reproducibly manufactured. Polynucleotides and chimeric compounds with acceptable toxicity profiles are particularly desirable.